Trial Information

Clinical and Biological Effects of 5-Azacitidine Five Days/Monthly Schedule in Symptomatic Low-risk Myelodysplastic Syndromes (MDSs)

5-Azacitidine(5-AZA) is the most promising drug for treatment MDSs. When administered at a
dose of 75mg/m2/day subcutaneously for 7 days, every 28 days (every month), 5-Aza produces
high rates of trilineage responses, reduces the risk of progression to AML in high-risk MDS
and improves the QoL. The use of 5-Aza in the earlier phases of MDS could reduce the
proliferative advantage of MDS clone and favour the regrowth of normal hematopoiesis. Other
doses or schedules could improve its efficacy. Gene expression profile studies in MDS
patients who are sensitive or resistant to 5-AZA are lacking and data coming from these
studies could be useful to clarify the mechanisms of 5-azacitidine and optimize the therapy.

Objectives of the study

Primary:

- To evaluate the efficacy (hematologic response) and toxicity of 5-Aza five days monthly
treatment schedule in patients with low-risk MDS (IPSS 0-INT1)

Secondary:

- To evaluate QoL by the FACT-An questionnaire

- To evaluate time to progression of MDS;

- To evaluate the gene expression profile of MDS patients sensitive or resistant to
5-AZA;

- To evaluate oncostatin M, interleukin-6 and interleukin-11 levels in the responders and
non responders.

Azacitidine will be given at a dose of 75mg/sqm subcutaneous daily for 5 consecutive days
every 28 days (every month) for a total of 8 courses. 5-Aza dosages will be adjusted as
follows:

Patients' care and other medications

- RBC are transfused to maintain a Hb level > 90 g/L, or whenever indicated.

- Platelets are transfused if platelet count is < 20 x 109/L, or whenever indicated.

- EPO is not allowed during the trial time (8 months= 8 courses of 5-AZA therapy)

- Granulocyte or granulocyte-monocyte colony stimulating factor (G-CSF or GM-CSF) are
allowed in case of severe neutropenia and/or systemic infection.

- Antibiotics and antifungals per os are given in prophylaxis if neutrophils <0.5 x 109
/L.

- Antibiotics are given i.v. in case of fever (> 38°C for > 24 h) or whenever indicated.

- Antifungals are given i.v. in case of fever persisting for more than 5 days of
antibiotics i.v. or whenever indicated.

- Other experimental drugs or agents are not allowed.

- If another experimental drug or agent will be administered, the patient must
discontinue the treatment with 5-AZA, and goes off-study.

Definition of Response

The response to treatment will be assessed according to IWG 2006 criteria, as reported by
Cheson et al.(28) (Appendix F)

Response criteria for altering natural history of MDS:

- Complete remission (CR) Bone marrow: ≤5% myeloblasts with normal maturation of all cell
lines Persistent dysplasia will be notes Peripheral blood: Hgb ≥ 11g/dl, Platelets ≥
100 x 109/l, Neutrophils ≥ 1.0 x 109/l, Blasts 0%

- Partial remission (PR)

All CR criteria if abnormal before treatment except:

Bone marrow blasts decreased by ≥50% over pretreatment but still >5% Cellularity and
morphology not relevant

- Marrow CR Bone marrow ≤ 5% myeloblasts and decrease by ≥ 50% over pretreatment
Peripheral blood: if HI response, they will be noted in addition to marrow CR

- Stable disease Failure to achieve at least PR, but no evidence of progression for >8
weeks

- Failure Death during treatment or disease progression characterized by worsening of
cytopenias, increase in the percentage bone marrow blasts, or progression to a more
advanced MDS FAB subtype than pretreatment.

- Relapse after CR or PR

At least one of the following:

- Return to pretreatment bone marrow blast percentage

- Decrement of ≥ 50% from maximum remission/response levels in granulocytes or platelets

- Reduction in Hgb concentration by ≥ 1.5 g/dl or transfusion dependence

- Cytogenetic Response

1. Complete Disappearance of the chromosomal abnormality without appearance of
new ones

2. Partial At least 50% reduction of the chromosomal abnormality

- Disease progression For patients with Less than 5% blasts: ≥50% increase in blasts
to > 5% blasts 5%-10% blasts: ≥50% increase to >10% blasts 10%-20% blasts: ≥ 50%
increase to > 20% blasts 20%-30% blasts: ≥50% increase to >30% blasts Any of the
following At least 50% decrement from maximum remission/response in granulocytes
or platelets Reduction in Hgb by ≥ 2g/dl Transfusion dependence

Survival

Endpoints:

Overall: death from any cause Event free: failure or death from any cause PFS: disease
progression or death form MDS DFS: time to relapse Cause-specific death: death related to
MDS

Response criteria for hematologic improvement (HI)

- Erythroid response (HI-E) (pretreatment, <11 g/dl) Hgb increase by ≥1.5 g/dl Relevant
reduction on units of RBC transfusion by an absolute number of at least 4 RBC
transfusions/8 weeks compared with the pretreatment transfusion number in the previous
8 weeks. Only RBC transfusions given for a Hgb of ≤ 9.0 g/dl pretreatment will count in
the RBC transfusion response evaluation

- Platelet response (HI-P)(pretreatment, <100 x 109/l) Absolute increase of ≥ 30 x 109/l
for patients starting with >20 x 109/l platelets Increase from < 20 x 109/l to >20 x
109/l and by at least 100%

- Neutrophil response (HI-N) (pretreatment, <1.0 x 109/l) At least 100% increase and an
absolute increase > 0.5 x 109/l

- Progression or relapse after HI

1. At least one of the following:

2. At least 50% decrement from maximum response levels in granulocytes or platelets

3. Reduction in Hgb by ≥ 1.5 g/dl Transfusion dependence

Quality of Life (QoL) QoL will be assessed by the FACT-An score before treatment and monthly
during the study, at the end of each cycle of treatment. The FACT-An questionnaire will be
performed as outpatient interviews by one trained nurse The QoL battery, according to
FACT-An questionnaire, consists of items including physical, functional, emotional, social
spiritual symptoms

Patient evaluation

- Physical examination (at entry and every month).

- Peripheral blood count and differential (at entry and every two weeks).

- Bone marrow biopsy (at entry; at the end of the 4th course; at the end of study-8th
course).

- Bone marrow aspirate (at entry; at the end of the 4th course; at the end of study-8th
course).

- Cytogenetics (at entry; at end of the 4th course; at the end of study -8th course).

- Blood biochemistry profile (at entry and every month).

- HBV, HCV, HIV (at entry).

- Gene Profile (at entry; at the end of the 4th course; at the end of study-8th course).

- Cytokines, Onc.M, IL6, IL11 (at entry; at the end of the 4th course; at the end of
study-8th course).

- FACT-an (at entry; at the end of each course).